Fate of an organic contaminant
Volatilization Leaching Sequestration Bioaccumulation Biodegradation
Bio – X - ation
Biodeterioration– BAD
Biodegradation– NEUTRAL
Bioremediation– GOOD
Microbial biodegradation
Aerobic Anaerobic In situ Ex situ Microbes + contaminants + TEA -> CO2 +
H2O + biomass
Requirements for life
Energy Water Carbon Nitrogen Oxygen? Phosphate Trace elements http://www.abe.iastate.edu/Ae573_ast475/
Stoichiometry_Notes.htm
Redox
In most cases the contaminant is oxidised (loses electrons). For this to happen, another compound needs to be reduced (gain electrons) to prevent electrons from accumulating. Usually there is a chain of these redox couples with the electrons eventually being taken up by a terminal electron acceptor
Oxygen CO2 Mn(IV) Mn(III) NO3
- NO2-
Fe(III) Fe(II) SO4
2- H2S H CH4
Hydrocarbon degradation
Aerobic Nitrate Manganese Fe(III) Fe(II) Sulphate Methanogenesis
Benzene
Toluene Ethylbenzene
m-Xylene p-Xylene o-Xylene Alkanes Alkenes LAB PAH Others
Redox zones
Vadose Zone
Saturated Zone
Bedrock
Water table
Aerobic
MethanogenicSulphate
Nitrate
Aerobic degradation ofn-alkanes
-oxidation Degrades hydrocarbon (fatty
acid) chain Removes 2 carbons at a time Ubiquitous pathway BUT needs O2
SH CoA
CH3
C97
S
O
CoA
OH2
SH CoA
CH2
112
CH
2
CH2
C115
O
O
R
RCH2
CH
2
CH2
C12
S
O
CoA
RCH2
CH
CH
S
O
CoA
RCH2
CH
CH2
C42
S
O
CoA
OH
RCH2
C56
CH2
C58
S
O
CoA
O
RCH2
C92
S
O
CoA
Oxidation
Hydration
Oxidation
Thiolysis
+
Activation
Fatty acid
Acyl CoA
Enoyl CoA
L-Hydroxyacyl CoA
Ketoacyl CoA
Acyl CoA Acetyl CoA
Beta oxidation (Adapted from Stryer 1981)
Initial anaerobic transformations of toluene
CH3
Toluene
CH3
OH
o-Cresol
CH3
OHp-Cresol
CH2OH
Benzyl Alcohol
CH3
Methylcyclohexane
Ring reduction/Ring cleavage/Mineralizationof Aliphatics
Anaerobic mineralizationof toluene
CH3
Toluene
CH2
CH
2
Hydrocinnamoyl-CoA
CH2
SCoA
O
O
Benzoyl-CoA
S
CoA
CoASH
CH2
CH C
H2
SCoA
O
B-hydroxycinnamoyl-CoA
OH
CH2
CH
CH
SCoA
O
Cinnamoyl-CoA
SCoA
O
CH2 CH2
SCoA
OO
B-ketocinnamoyl-CoA
CO2
OH2
SCoA
O
Proposed pathway for anaerobic toluene mineralization - after Chee-Sanford et al 1996
2e-, 2H+ 2e-, 2H+
2e-, 2H+
Anaerobic degradation of toluene
CH3
Toluene
CH2
CH
Benzyl-succinate
CH2
OO
O
O
O
Benzoyl-CoA
S
CoA
CoASH Succinyl-CoA
CH CH2
O
O
O
E-Phenylitaconyl-CoA
S
CoA
CH C
H CH2
O
O
O
2-Carboxymethyl-3-Hydroxy-Phenylpropionyl-CoA
OH
S
CoA
OH2
CH C
H2
O
O
O
O
S
CoA
Benzylsuccinyl-CoA
CH2
CH C
H2
O
O
OS
CoA
Benzylsuccinyl CoA
BenzylsuccinateSynthetase
Benzylsuccinate-CoA Transferase Benzylsuccinyl-CoA
Dehydrogenase
3-Hydroxyacyl-CoADehydrogenase
Benzoylacetyl-CoAThiolaseolase
Proposed pathway for anaerobic toluene degradation - after Heider et al 1999
FumarateSuccinyl CoA Succinate
Phenylitaconyl-CoAHydratase
2[H]
2[H]
Anaerobic ethylbenzene degradation
CH2
CH3
Ethylbenzene
CH CH3OH
1-Phenylethanol
CH3O
Acetophenone
O O
CH2O
Benzoylacetate
O O
CH2O
SCoA
Benzoylacetate-CoA
O S
CoA
Benzoyl-CoA
OH2 CO2 CoASH CoASH Acetyl-CoA
EthylbenzeneDehydrogenase
1-PhenylethanolDehydrogenase
AcetophenoneCarboxylase
Benzoylacetyl-CoAforming enzyme
Benzoylacetyl-CoACoA thiolase
Proposed pathway for anaerobic ethylbenzene degradation - after Heider et al 1999
2[H]2[H]
Anaerobic alkylbenzene degradation
Toluenem-Xylene
p-Xylene
Ethylbenzeneo-Xylene
COOH
COOH
COOH
COOH
CH3
COOH
COOH
CH3
COOH
COOH
CH3
COOH
COOH
CH3
CH3
COOH COOH
CH3
COOH
CH3
CH3
COOHCOOH
CH3
COOH
CH3
COOH
COOH
COOH
COOH
Elshahed et al 2001
Alkylbenzenes (T,E,X) Key metabolites in
degradation All seen in
laboratory/ground water
Benzoate
Anaerobic benzoate degradation
SCoAO SCoAO
SCoAO SCoAO
OH
SCoAO
OCOO-
SCoAO
COO-
SCoAO
COO-
SCoAO
OH
SCoAO
OH
SCoAO
OH OH
SCoAO
OH O
OHO
B e n z o y l - C o A
P i m e l y l - C o A
C y c l o h e x - 1 , 5 - d i e n e1 - c a r b o x y l - C o A
6 - H y d r o x y c y c l o h e x - 2 - e n e -1 - c a r b o x y l - C o A
C y c l o h e x - 1 - e n e1 - c a r b o x y l - C o A
2 - H y d r o x y c y c l o h e x a n e -1 - c a r b o x y l - C o A
2 - K e t o C y c l o h e x a n e -1 - c a r b o x y l - C o A
2 , 3 - D e d e h y d r o -p i m e l y l - C o A
3 - H y d r o x y p i m e l y l - C o A
2 , 6 - D i h y d r o x y c y c l o h e x a n e -1 - c a r b o x y l - C o A
6 - O x o - 2 - h y d r o x y c y c l o h e x a n e -1 - c a r b o x y l - C o A
E1
E2E3 E4 E5 E6
E7
E8
E9 E10
E11
E1 -- Benzoyl-CoA reductase
E2 -- Cyclohex-1,5-diene -carboxyl-CoA reductase
E3 -- Cyclohex-1-ene 1-carboxyl-CoA hydratase
E4 -- 2-Hydroxycyclohexane-1-carboxyl-CoA dehydrogenase
E5 -- 2-Ketocyclohexane11-carboxyl-CoA hydrolase
E6 -- Pimelyl-CoA dehydrogenase
E7 -- 3-hydroxyacyl-CoA deyhdratase
E8 -- Cyclohex-1,5-diene-1-carboxyl-CoA hydratase
E9 -- 6-Hydroxycyclohex-2-ene-1-carboxyl-CoA hydratase
E10 -- 2,6-Dihydroxycyclohexane-1-carboxyl-CoA dehydrogenase
E11 -- 6-Oxo-2-hydroxycyclohexane-1-carboxyl-CoA hydrolase
Harwood and Gibson (1997) and Koch et al. (1993)
B e n z o a t e
Anaerobic degradation of n-alkanes
Limited range of chain lengths No < 6 C to date Pathways unknown Specific to organism May involve addition/removal of
odd number of C Rate of dissolution may limit rate
of degradation
Aerobic degradation of LAB
If chain > 3 long then starts with -oxidation of methyl terminus/i
Ring cleavage by oxidationR
CH
CH
R
OH
OH
R
OH
OH
R
OH
COOHO
O
COOH
RCOOH
NAD+ NADHNADH NAD+
O2 O2
+
Alkylbenzene Dihydrodiol Ring fissionproduct 2-Oxopenta-
4-enoate
2,3-Dihydroxy-alkylbenzene
Smith & Ratledge 1989
E2 E3 E4E1
E1 = Alkylbenzene dioxygenase
E2 = cis-alkylbenzene glycol dehydrogenase
E3 = 2,3-dihydroxyalkylbenzene 1,2-dioxygenase
E4 = ring fission product-hydrolysing enzyme
Anaerobic degradation of LAB
-oxidation? Conversion to benzoyl CoA? Hydrolytic ring cleavage? Limited by rate of dissolution?
Generalized breakdown
HYDROCARBON (eg BTEX)
Aerobic
Chain degraded by Beta oxidation
Convert to e.g.benzoyl CoA
Ring cleavage by hydrolysis (add H2O)
Anaerobic
?
Ring cleavage by oxygenases (add O2)
Organisms
Bacteria– bioremediation
Fungi– mycoremediation
Plants– phytoremediation
Bioavailability
May not be available to organisms Chemically Physically
– Solubility– Sorption
Pathways
The University of Minnesota Biocatalysis/Biodegradation Database– http://umbbd.ahc.umn.edu/
Bioremediation techniques
MNA/MENA Landfarming Bioventing/sparging Windrows Composting Biopile Biofiltration Bioaugmentation Biostimulation Redox/TEA
Group 1Not degraded
Bitumen Asphalt Metals Inorganic acids Asbestos Complex cyanides
Group 2May be degraded in lab
Higher MW PAHs PCBs Tars
Group 3 Demonstrated but not regularly achieved
Explosives Pesticides (e.g. lindane, malathion, diuron,
mecoprop, paraquat) PCP High MW PAH Branching aliphatics (e.g. hopane) Surfactants (e.g.LAS) MTBE Complex cyanides (?)
Group 4Regularly treated aerobically
Diesel Jet fuel BTEX Paraffin Ammonia Crude oil Lubricating oil Petrol Phenol
Chlorophenols Organic acids Creosote Alcohols Aldehydes Ketones Some surfactants Some pesticides Low MW PAHs
Group 5Regularly treated anaerobically
Chlorinated solvents
Soil factors affecting degradation
Organic matter content Microbial activity pH
– ionisable compounds– Acid/base catalysed degradation
Temperature Soil water content Depth
– Faster near surface
Phytoremediation
Rhizoremediation Transpiration (control flow) Volatilisation Plant metabolism Accumulation Stabilisation/immobilisation